PROJECT SUMMARY In the United States, musculoskeletal diseases such as extremity injuries, burns, and tumors are a leading cause of disabilities and death, affecting one in two individuals. However, until now, there has been no effective implant that can replace the structure and function of damaged bone and muscle tissues, likely due to the difficulty of regulating the sophisticated heterogeneous bone-muscle junction structure. As a result, muscle damage has been largely ignored during musculoskeletal surgeries, which often results in disconnected tissues and fibrous tissue formation, leading to temporal or permanent musculoskeletal disability. In fact, in the human musculoskeletal system, there exists a direct attachment between bone and muscle tissues at a wide area of bone, forming a ?bone-muscle unit.? Based on this structural closeness, the growth and development of bone and muscle are tightly coupled through growth factor signaling and cellular cross-talk. Therefore, damage to either bone or muscle can deteriorate health and function of the other tissue type. For this reason, there has been a strong need for developing an innovative musculoskeletal implant, which can integrate the distinguished physicochemical properties of hard tissue and soft tissue in a spatially controlled manner. To address this problem, we aim to design and build the first 3D printed muscle-bone implant, by utilizing state- of-the-art 3D multimaterial bioprinting that can extrude multiple types of tissue mimetic bioinks in a simultaneous and continuous manner. We will control the physicochemical properties of bioinks, such as viscosity and porosity, to provide an optimized artificial niche for the growth and differentiation of each cell type. We will also include biodegradable drug carriers to supply musculogenic and osteogenic growth factors with controlled release kinetic behavior, to aid tissue recovery. In addition, we will regulate the parameters for bioprinting, such as pneumatic pressure, and the injection and photocrosslinking conditions to build a 3D structure. We will then mature the 3D printed muscle-bone organ implant in a customized bioreactor system by applying compression and relaxation cycles that mimic musculoskeletal movement in vivo. Finally, we will evaluate the musculoskeletal regeneration capacity of our 3D printed muscle-bone implant in a mouse volumetric muscle loss and bone defect model. This research will present the first 3D print muscle-bone tissues with continuous structures ex vivo that can provide a groundbreaking clinical solution for curing severe musculoskeletal injuries and preventing disabilities in the clinic. We further expect that our 3D printed muscle-bone tissue platform will be beneficial for understanding developmental principles and pathological mechanisms of the musculoskeletal system.